Antiphospholipid antibodies occur in autoimmune diseases such as systemic lupus erythematosus (SLE) and antiphospholipid antibody syndrome (APS) as well as in association with infections and drug therapy. APS is characterized by one or more clinical features such as arterial or venous thrombosis, thrombocytopenia and fetal loss. APS may be primary or it may be associated with other conditions, primarily SLE. (PHOSPHOLIPID-BINDING ANTIBODIES (Harris et al., eds., CRC Press, Boca Raton, Fla., 1991) ; McNeil et al. ADVANCES IN IMMUNOLOGY, Vol. 49, pp. 193-281 (Austen et al., eds., Academic Press, San Diego, Calif., 1991)). Approximately 30-40% of patients with SLE have aPL, however, 50% of patients with aPL antibodies do not have SLE. This 50% may have other autoimmune rheumatic diseases, miscellaneous conditions or they may have been subjected to drug therapy, particularly chlorpromazine. In one study of 70 patients, 26 males and 44 females, with primary APS (PAPS) but no evidence of SLE, the following features were observed: deep venous thrombosis (DVT) in 31; arterial occlusion in 31, particularly stroke or transient ischemia; myocardial infarctions in 15; recurrent fetal loss in 24; thrombocytopenia (TCP) in 32; 10 had a positive Coombs' test; Evans' syndrome in 7; anti-nuclear antibody (ANA) in 32, but less than 1:160 in 29; and antimitochondrial antibody (AMA) in approximately 24. (McNeil et al., supra.). Estimates vary but in about 5% of all stroke patients, aPL antibodies are thought to be an important contributing factor.
Transient aPL antibodies, such as those detected in a VDRL test, occur during many infections. Approximately 30% of patients possessing persistent aPL antibodies have suffered thrombic event. The presence of aPL antibodies defines a group of patients within SLE who display a syndrome of clinical features consisting of one or more of thrombosis, TCP, and fetal loss. The risk of this syndrome in SLE overall is around 25%; this risk increases to 40% in the presence of aPL antibodies and decreases to 15% in their absence. Because aPL antibodies appear to be directed at phospholipids in plasma membranes, it has been postulated that they may exert direct pathogenic effects in vivo by interfering with hemostatic processes that take place on the phospholipid membranes of cells such as platelets or endothelium. In patients with PAPS, the fact that aPL antibodies appear to be the only risk factor present is further evidence that these antibodies have a direct pathogenic role. Induction of PAPS by immunizing mice with human anticardiolipin antibodies is the best evidence yet that aPL antibodies are directly pathogenic (Bakimer et al. 1992 J. Clin. Invest. 89:1558-1563; Blank et al. 1991 Proc. Natl. Acad. Sci. 88:3069-3073).
It is now generally accepted that aPL antibodies recognize an antigenic complex comprised of .beta..sub.2 -glycoprotein I (.beta..sub.2 -GPI) and negatively-charged phospholipid, e.g., cardiolipin (McNeil et al. (1990) Proc. Natl. Acad. Sci. 87:4120-4124; Galli et al. (1990) Lancet i:1544-1547). .beta..sub.2 -GPI is a minor plasma glycoprotein found free and in association with lipoprotein lipids where it is also known as apolipoprotein H (apo H). It consists of five independently folding domains referred to as Sushi or short consensus repeat domains that resemble similar domains in other proteins. .beta..sub.2 -GPI has been reported to undergo antigenic and conformational changes upon binding phospholipid (Wagenkneckt et al. (1993) Thromb. Haemostas. 69:361-365; Jones et al. (1992) Proc. 5th Intl. Symp. Antiphospholipid Antibodies (Abstract S5)). The fifth domain of .beta..sub.2 -GPI is the site of lipid binding as well as the site recognized by aPL antibodies (Hunt J. and S. Krilis, (1994) J. Immunol. 152:653-659; Lauer et al. (1993) Immunol. 80:22-28). The pathological mechanism for aPL is unknown (McNeil et al., supra). Most explanations invoke endothelial cell function or platelet involvement (Haselaar et al. (1990) Thromb. Haemostas. 63:169-173). These explanations suggest that following blood vessel endothelial cell injury or platelet activation, the exposure or transbilayer migration of anionic phospholipid to the plasma-exposed surface may lead to .beta..sub.2 -GPI-binding and trigger aPL antibody formation.
aPL antibodies may be directly prothrombotic by reducing prostacyclin formation (Vermylen, J. and J. Arnout (1992) J. Clin. Lab. Med. 120:10-12); by direct interference with the action of coagulation proteins; or by blocking the ability of .beta..sub.2 -GPI to inhibit the intrinsic blood coagulation pathway, platelet prothrombinase activity, and ADP-mediated platelet aggregation (Arvieux et al. (1993) Thromb. Haemostas. 60:336-341).
A major new tool in medicinal chemistry in the search for lead compounds has been the advent of combinatorial libraries providing vast molecular diversity. Molecular diversity may arise from chemical synthesis or from biological systems (Scott., J. K. RATIONAL DRUG DESIGN (CRC Press, Weiner, D. B. and W. V. Williams, eds., Boca, Raton, Fla., 1994); Moos et al. (1993) Ann. Reports Med. Chem. 28:315-324). By displaying random peptides on the surface of filamentous phage, epitope libraries containing hundreds of millions of clones for probing by clinically significant antibodies have been created (Scott, J. K. and G. P. Smith (1990) Science 249:286-390; Cesareni, G. (1992) FEBS Lett. 307:66-70). Such phage libraries are prepared by incorporating randomized oligonucleotide sequences into the phage genome, usually the pIII gene, which encode unique peptide sequences on the surface of each phage. Following sequential rounds of affinity purification and amplification, those phage that bind antibody are propagated in E. coli and the binding peptides identified by sequencing the corresponding coding region of viral DNA. In most cases, subsequent study will involve corresponding synthetic peptides after establishing their ability to bind antibody. Phage-based libraries have been used to mimic discontinuous epitopes (Luzzago et al. (1993) Gene 128:51-57; BaLass et al. (1993) Proc. Natl. Acad. Sci. 90:10638-10642). The potential plasma instability of peptide-based drugs has been successfully overcome by N-terminal blocking or by the judicious use of amino acid analogs (Powell, M. F. (1993) Ann. Reports Med. Chem. 28:285-293).
At present there is no selective, immunospecific therapy for patients showing high titers of aPL antibodies. In many cases use of drugs such as aspirin, steroids, and warfarin has proven to be largely inadequate (PHOSPHOLIPID-BINDING ANTIBODIES (Harris et al., eds., CRC Press, Boca Raton, Fla., 1991); McNeil et al., supra)). Synthetic mimotope peptides, characterized by (i) the inability to activate T cells while (ii) retaining the ability to bind immune B cells, are used to tolerize in an antigen-specific manner. This technology is disclosed in co-owned, co-pending U.S. patent application, Ser. No. 08/118,055, filed Sep. 8, 1993, and U.S. Pat. No. 5,268,454, which are incorporated by reference herein in their entirety. As disclosed in the application and patent cited above, B cell tolerance entails administering such peptides conjugated to multivalent, stable, non-immunogenic valency platforms in order to abrogate antibody production via B cell anergy or clonal deletion after cross-linking surface immunoglobulin.
Although the exact molecular nature of the target epitopes recognized by aPL antibodies is unknown, the use of peptides derived from epitope libraries will allow for the construction of successful tolerogens. B cell tolerogens for the treatment of human systemic lupus erythematosus-related nephritis have also been disclosed in co-owned U.S. Pat. Nos. 5,276,013 and 5,162,515 which are incorporated by reference herein in their entirety.